Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0017636 (glioblastoma)
 18,345 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Recent cloning studies confirm the presence of two subtypes of bombesin (Bn) receptors. In contrast to the gastrin-releasing peptide (GRP)-preferring subtype, which has been widely studied, nothing is known about the cellular mechanisms of the neuromedin B (NMB)-preferring subtype, which occurs widely in the central nervous system and gastrointestinal tissues, partially because of the lack of a cell line with functional receptors. In the present study we have investigated Bn receptors on the rat glioblastoma cell line C-6, reported to contain mRNA of the NMB receptor subtype. Binding of 125I-[D-Tyr0]NMB to these cells was time- and temperature-dependent, saturable, reversible, and only inhibited by Bn receptor agonists or antagonists. For Bn receptor agonists the relative potencies were: NMB (1.7 nM) approximately equal to litorin (3 nM) greater than ranatensin (8 nM) greater than Bn (19 nM) greater than neuromedin C (NMC) (210 nM) greater than GRP (500 nM). These relative affinities were almost identical to those for the NMB receptor subtype on rat oesophageal tissue and for Balb 3T3 cells stably transfected with the NMB receptor subtype. These potencies differed from those for the GRP receptor subtype on rat pancreatic acini [Bn approximately equal to litorin (4 nM) greater than ranatensin, NMC, GRP (15-20 nM) much greater than NMB (351 nM)]. The relative potencies of four different classes of Bn receptor antagonists were compared. Results from C-6 tumour cells agreed closely with those for binding to the NMB receptor subtype on rat oesophageal tissue and in Balb 3T3 cells stably transfected with this receptor, and differed markedly from those for binding to the GRP receptor subtype on rat pancreatic acini. Four Bn receptor antagonists had a higher affinity for the GRP subtype ([D-Phe6]Bn-(6-13)ethyl ester (500 x), [D-Phe6][psi 13-14,Cpa14]Bn- (6-14) (70 x) (where psi 13-14 refers to the replacement of the -CONH- peptide bond between Leu13 and Met14 by -CH2NH2) [psi 13-14,Leu14]Bn, [D-Phe6]Bn-(6-13) propylamide (30 x)] and two had a higher affinity for the NMB subtype on C-6 cells and transfected cells ([D-Pro4,D-Trp7,9,10] substance P-(4-11) (9 x) and [Tyr4,D-Phe12]Bn (18 x)]. In C-6 tumour cells, Bn receptor agonists caused an increase in cytosolic Ca2+ and the generation of inositol phosphates. For both responses, NMB was more than 50-fold more potent than GRP. Neither NMB nor GRP increased cyclic AMP. These results demonstrate that the rat glioblastoma cell line C-6 possesses functional NMB-preferring Bn receptors, and agonist occupation activates phospholipase C, thus increasing cytosolic Ca2+ and inositol phosphate formation. Because the interaction of Bn-related peptides with C-6 cell receptors is identical with that reported in other tissues containing the mRNA for the NMB subtype, this cell line should prove useful in exploring further the cellular basis of action of the peptides that interact with this receptor in the central nervous system and various other tissues.
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PMID:Activation of neuromedin B-preferring bombesin receptors on rat glioblastoma C-6 cells increases cellular Ca2+ and phosphoinositides. 132 46

The receptor that interacts with the mammalian bombesin-related peptide neuromedin B (NMB) is ubiquitous in the gastrointestinal tract and central nervous system. However, little is known regarding its cellular mechanisms of action. This receptor has been recently cloned, sequenced, and stably transfected into BALB 3T3 fibroblasts, permitting detailed study of the pharmacology and coupled biological activities of this receptor. In the present study, we compare the ability of transfected receptors to alter cell function with that of receptors natively expressed in small numbers by the rat glioblastoma cell line C6. NMB inhibited binding of 125I-[D-Tyro]NMB with high affinity in transfected cells (Ki = 3.08 +/- 0.14 nM) and in C6 cells (Ki = 1.90 +/- 1.10 nM), whereas the bombesin-related agonists gastrin-releasing peptide (GRP) and [D-Phe6, D-Ala11, Leu14]bombesin(6-16) (GRP analogue) had 100- and 300-fold lower affinities, respectively, for NMB receptors in either cell type. For both cell systems, maximal binding was observed between 5 and 15 min at 22 degrees. Both cell types internalized NMB at similar rates, with > 70% of bound ligand being internalized by 60 min at 22 degrees. The nonhydrolyzable guanosine analogue guanosine 5'-(beta,gamma-imido)triphosphate was equipotent in causing a decrease in binding of 125I-[D-Tyro]NMB due to decreased receptor affinity in both cell types, without a change in receptor number, demonstrating that the NMB receptor remained coupled to a guanine nucleotide-binding protein in both native and transfected cells. In both cell systems, NMB increased inositol monophosphate, inositol bisphosphate, and inositol trisphosphate in a time-dependent fashion. Inositol phosphates were increased in a dose-dependent fashion, with similar half-maximal values being obtained for NMB in both cell types (transfected, 1.01 +/- 0.09 nM; C6, 2.09 +/- 0.15 nM) and for the GRP analogue (transfected, 1855 +/- 140 nM; C6, 2129 +/- 250 nM). NMB mobilized intracellular Ca2+ in both cell systems, and the dose-response curves were superimposible (EC50 for transfected, 0.10 +/- 0.08 nM; C6, 0.11 +/- 0.02 nM). These data demonstrate that activation of the receptor for NMB stimulates phospholipase C and increases intracellular Ca2+. These results also demonstrate that transfected and native NMB receptors behave similarly, suggesting that the transfected cell line will be useful in future studies investigating ligand-receptor interactions, as well as in molecular biological studies of the structure-function relationship of the receptor.
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PMID:Neuromedin B receptors retain functional expression when transfected into BALB 3T3 fibroblasts: analysis of binding, kinetics, stoichiometry, modulation by guanine nucleotide-binding proteins, and signal transduction and comparison with natively expressed receptors. 133 12

Little is known about the pharmacology or cell biology of human bombesin (Bn) receptors, because they are usually present at low levels and both subtypes are frequently present in the same tissues. Human gastrin-releasing peptide (GRP) receptors (huGRP-R) and human neuromedin B (NMB) receptors (huNMB-R) were stably transfected into BALB/3T3 fibroblasts. Both receptor types were glycosylated, with 35% of the huGRP-R and 38% of the huNMB-R representing carbohydrate residues. The extent of glycosylation of the transfected huGRP-R was the same as that seen in the human glioblastoma cell line U-118. Radiolabeled agonist ligands were rapidly internalized, whereas noninternalized ligand readily dissociated in a temperature-dependent fashion. The affinities of various agonists for binding to the huGRP-R were Bn (Ki = 1.4 +/- 0.2 nM) = 4 x GRP = 300 x NMB. In contrast, affinities for the huNMB-R were NMB (Ki = 8.1 +/- 5.2 nM) = 4 x Bn = 600 x GRP. [F5-D-Phe6,D-Ala11]Bn(6-13)methyl ester was the most potent huGRP-R antagonist, whereas D-Nal-Cys-Tyr-D-Trp-Lys-Val-Cys-Nal-NH2 was the most potent huNMB-R antagonist. Agonist binding to either receptor type caused activation of phospholipase C and increased cellular [3H]inositol phosphate levels. GRP was potent at increasing [3H]inositol phosphate generation in cells expressing the huGRP-R (EC50 = 13.6 +/- 1.3 nM), whereas NMB was similarly potent when acting upon cells expressing the huNMB-R (EC50 = 9.3 +/- 1.4 nM). However, neither receptor type, when stimulated with agonist, caused an increase in cAMP levels. These data show that stably transfected huGRP-R exhibit similar pharmacology for agonists and antagonists, are appropriately glycosylated, and function similarly with respect to their ability to alter biological activity, compared with natively expressed receptors. Minimal native huNMB-R data are available for comparison, but in general the huNMB-R is similar to the rat NMB receptor in its pharmacology and cell biology.
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PMID:Expression and characterization of cloned human bombesin receptors. 783 18

L-Proline transport in C6 glioblastoma cells takes place mainly via a saturable Na(+)-dependent mechanism. The uptake process can be discriminated into two components, system A and system ASC. A minor proportion of L-proline transport is carried out by the ASC system, which appears to be constitutively expressed by the cell, but most is by system A which shows adaptive responses to amino acid deprivation and sensitivity to N-methyl-alpha-aminoisobutyric acid. The transport system is inhibited by proline derivatives, such as methyl and benzyl esters, and also hydroxyproline, and is stereospecific. Incubation of glioblastoma cells with phorbol 12-myristate 13-acetate led to concentration- and time-dependent decreases in L-proline transport. This effect could be mimicked by exogenous phospholipase C. Proline transport is significantly stimulated in the presence of Ca(2+)-mobilization agents and strongly inhibited in the absence of Ca2+. The present data suggest a complex regulation of L-proline transport by different kinases in glioblastoma cells.
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PMID:Characteristics and regulation of proline transport in cultured glioblastoma cells. 794 91

The receptor for neuromedin B (NMB-R), a mammalian bombesin-related peptide, is widely distributed in the central nervous system and gastrointestinal tract. While it is known that this receptor is coupled to phospholipase C, like many other phospholipase C-activating receptors, little is known about regulation of the NMB-R subsequent to agonist stimulation. We studied both native NMB-R on C-6 rat glioblastoma cells and wild type NMB-R cloned from rat esophageal muscle which was stably transfected into Balb/3T3 fibroblasts. Both cell types rapidly increased [3H]inositol phosphates and [Ca2+]i in response to 1 microM NMB, whereas preincubation with 3 nM NMB for 3 h markedly attenuated the ability of 1 microM NMB, but not 1 microM endothelin-1, to alter either cell type's biological activity. Prolonged exposure to 3 nM NMB caused a rapid decrease in the number of NMB-R, with the maximal receptor down-regulation seen at 24 h due to NMB-R internalization. After maximal down-regulation, removal of agonist resulted in a rapid restoration of NMB-R to the cell surface of both cell types. NMB-R recovery at 6 h was blocked by monensin, an inhibitor of receptor recycling, but was not affected by cycloheximide, a protein synthesis inhibitor. Resensitization to agonist paralleled the recovery of NMB-R in both cell types, and resensitization likewise was blocked by monensin. Our data demonstrate that the NMB-R undergoes rapid homologous desensitization consequent to agonist stimulation, which is mediated by receptor down-regulation and which, in turn, is regulated by internalization. During resensitization, NMB-R reappearance on the cell surface membrane is independent of protein synthesis and is due to a recycling from an intracellular site.
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PMID:Desensitization of neuromedin B receptors (NMB-R) on native and NMB-R-transfected cells involves down-regulation and internalization. 816 69

The actions of neuromedin B (NMB), a recently discovered mammalian bombesin-related peptide, are mediated by interacting with a distinct receptor; however, little is known about its cellular basis of action. Recent studies show activation of phospholipase D (PLD) is an important transduction cascade for a number of GI hormones, especially for stimulation of growth and protein sorting. The purpose of the present study was to determine whether activation of the NMB receptor causes activation of PLD and to explore whether this activation was coupled to PLC activation. Rat C6 glioblastoma cells (C6 cells), which contain a low density of native NMB receptors and BALB 3T3 cells stably transfected with rat NMB receptors, were used. NMB caused a 3-fold increase in C6 cells and an 11-fold increase in rNMB-R transfected cells in PLD activity. Increases in PLD activity were rapid and NMB was 100-fold more potent than gastrin-releasing peptide (GRP). NMB caused a half-maximal increase in [Ca2+]i at 0.2 nM, in [3H]IP and PLD at 1 nM, and half-maximal receptor occupation at 1.2 nM. TPA increased PLD dose-dependently with a half-maximal effect at 60 nM. The calcium ionophore A23187 (1 microM) alone did not increase PLD activity but potentiated the effect of TPA. The Ca2+-ATPase inhibitor, thapsigargin, did not affect NMB- or TPA-stimulated PLD activities, although it blocked completely the NMB-induced increase in [Ca2+]i. The PKC inhibitor GF109203X completely abolished TPA-induced PLD activity, however, it only inhibited NMB-induced PLD activity by 20%. The combination of thapsigargin and GF109203X had the same effect as GF109203X alone. These data indicate that NMB receptor activation is coupled to both PLC and PLD. In contrast to a number of other phospholipase C-coupled receptors, NMB receptor stimulated changes in [Ca2+]i do not contribute to PLD activation. Both PKC-dependent and PKC-independent mechanisms are involved in the NMB-stimulated PLD activation with the PKC-independent pathway predominating.
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PMID:Neuromedin B activates phospholipase D through both PKC-dependent and PKC-independent mechanisms. 955 86

Glioblastoma multiforme is a malignant primary brain tumor associated with short patient survival in part because of the ability of individual cells to migrate significant distances into brain tissue. Invasion is a difficult process to model, because many such human tumors do not invade immunologically competent animal tissue, tumors grown in animals do not invade human tissue, and relevant human tissue substrates are not easily reproduced. We discuss models for examining invasion in vitro, and in particular review work using the tumor spheroid--fetal rat brain aggregate co-culture model, assessed with confocal microscopy and four-dimensional imaging. Quantitation of invasion in this model is discussed, as well as the invasion-inhibitory properties of tyrosine kinase (TK) inhibitors. The effects of receptor-specific tyrphostins strongly support a dominant role for Epidermal Growth Factor Receptor activation in this process and show that invasion can be effectively inhibited at much lower concentrations of TK inhibitors than is necessary for growth suppression. Inhibition of activation of the purported growth factor receptor second messenger phospholipase C- gamma 1, by pharmacological means and gene transfection, also profoundly inhibits the invasive properties of human glioblastoma and rat C6 glioma cells. We have assessed invasiveness in several human tumor specimens, which may provide information relative to prognosis and recurrence risk. Our data supports the concept of differential control of invasion and proliferation, and points to possible strategies for anti-invasive therapy for glioblastoma multiforme.
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PMID:Inhibition of glioma invasion of fetal brain aggregates. 957 29

Enhanced activity of receptor tyrosine kinases such as the PDGF beta-receptor and EGF receptor has been implicated as a contributing factor in the development of malignant and nonmalignant proliferative diseases such as cancer and atherosclerosis. Several epidemiological studies suggest that green tea may prevent the development of cancer and atherosclerosis. One of the major constituents of green tea is the polyphenol epigallocathechin-3 gallate (EGCG). In an attempt to offer a possible explanation for the anti-cancer and anti-atherosclerotic activity of EGCG, we examined the effect of EGCG on the PDGF-BB-, EGF-, angiotensin II-, and FCS-induced activation of the 44 kDa and 42 kDa mitogen-activated protein (MAP) kinase isoforms (p44(mapk)/p42(mapk)) in cultured vascular smooth muscle cells (VSMCs) from rat aorta. VSMCs were treated with EGCG (1-100 microM) for 24 h and stimulated with the above mentioned agonists for different time periods. Stimulation of the p44(mapk)/p42(mapk) was detected by the enhanced Western blotting method using phospho-specific MAP kinase antibodies that recognized the Tyr204-phosphorylated (active) isoforms. Treatment of VSMCs with 10 and 50 microM EGCG resulted in an 80% and a complete inhibition of the PDGF-BB-induced activation of MAP kinase isoforms, respectively. In striking contrast, EGCG (1-100 microM) did not influence MAP kinase activation by EGF, angiotensin II, and FCS. Similarly, the maximal effect of PDGF-BB on the c-fos and egr-1 mRNA expression as well as on intracellular free Ca2+ concentration was completely inhibited in EGCG-treated VSMCs, whereas the effect of EGF was not affected. Quantification of the immunoprecipitated tyrosine-phosphorylated PDGF-Rbeta, phosphatidylinositol 3'-kinase, and phospholipase C-gamma1 by the enhanced Western blotting method revealed that EGCG treatment effectively inhibits tyrosine phosphorylation of these kinases in VSMCs. Furthermore, we show that spheroid formation of human glioblastoma cells (A172) and colony formation of sis-transfected NIH 3T3 cells in semisolid agar are completely inhibited by 20-50 microM EGCG. Our findings demonstrate that EGCG is a selective inhibitor of the tyrosine phosphorylation of PDGF-Rbeta and its downstream signaling pathway. The present findings may partly explain the anti-cancer and anti-atherosclerotic activity of green tea.
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PMID:Epigallocathechin-3 gallate selectively inhibits the PDGF-BB-induced intracellular signaling transduction pathway in vascular smooth muscle cells and inhibits transformation of sis-transfected NIH 3T3 fibroblasts and human glioblastoma cells (A172). 1019 59

The effect of the green tea compounds 2-(3,4-dihydroxyphenyl)-3, 4-dihydro-2H-1-benzopyran-3,5,7-triol (catechin), epicathechin (EC), epigallocathechin-3 gallate (EGCG), epicathechin-3 gallate (ECG) and catechin-3 gallate (CG) on the tyrosine phosphorylation of PDGF beta-receptor (PDGF-Rbeta) and on the anchorage-independent growth of A172 glioblastoma cells in semisolid agar has been investigated. Treatment of A172 glioblastoma with 50 microM CG, ECG, EGCG and 25 microM Tyrphostin 1296 resulted in an 82+/-17%, 77+/-21%, 75+/-8% and 55+/-11%, respectively (mean+/-S.D., n=3) inhibition of the PDGF-BB-induced tyrosine phosphorylation of PDGF-Rbeta. The PDGF-Rbeta downstream intracellular transduction pathway including tyrosine phosphorylation of phospholipase C-gamma1 (PLC-gamma1) and phosphatidylinositol 3'-kinase (PI 3'-K) was also inhibited. Spheroid formation was completely inhibited by 50 microM ECG, CG, EGCG and by 25 microM Tyrphostin 1296. We conclude that catechins of the green tea possessing the gallate group in their chemical structure act as anticancer agents probably partly via their ability to suppress the tyrosine kinase activity of the PDGF-Rbeta.
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PMID:Green tea compounds inhibit tyrosine phosphorylation of PDGF beta-receptor and transformation of A172 human glioblastoma. 1076 May 11

Neuropeptide Y (NPY) regulates neurotransmitter release through activation of the Y2 receptor subtype. We have recently characterized a human glioblastoma cell line, LN319, that expresses exclusively NPY Y2 receptors and have demonstrated that NPY triggers transient decreases in cAMP and increases in intracellular calcium responses. The present study was designed to further characterize calcium signalling by NPY and bradykinin (BK) in LN319 cells. Both agonists elevated free intracellular calcium ([Ca(2+)](i)) without soliciting calcium influx. NPY appeared to activate two distinct signalling cascades that liberate calcium from thapsigargin- and ryanodine-insensitive compartments. One pathway proceeded through phospholipase C (PLC)-dependent phosphatidylinositol turnover, while the other triggered calcium release through a so far unidentified mediator. Part of the response was sensitive to pertussis toxin (PTX) under conditions where the toxin totally abolished the NPY-mediated effects on cAMP. The calcium release induced by BK on the other hand was largely PTX-insensitive, PLC-dependent, and from both thapsigargin- and ryanodine-sensitive stores. Following stimulation with NPY, subsequent [Ca(2+)](i) responses to NPY were strongly depressed. Partial heterologous desensitization occurred, when BK was used as the first agonist, whereas NPY had no effect on a subsequent stimulation with BK. These data suggest that NPY-induced calcium mobilization in LN319 cells involves two different G proteins and signalling mediators, and a hitherto unidentified calcium compartment. Homologous desensitization of NPY signalling might be explained by receptor-G protein uncoupling, while heterologous desensitization by BK could be the result of either transient depletion or inhibition of a mediator in the calcium signalling cascades activated by NPY.
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PMID:Neuropeptide Y Y2 receptor signalling mechanisms in the human glioblastoma cell line LN319. 1128 92


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